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Alkaline phosphatase (AP) is a widely distributed non-specific
phosphomonoesterase that functions through formation of a covalent phosphoseryl
intermediate (E-P). The enzyme also catalyzes phosphoryl transfer reaction to
various alcohols. Escherichia coli AP is a homodimer with 449 residues per
monomer. It is a metalloenzyme with two Zn2+ and one Mg2+ at each active site.
The crystal structure of native E. coli AP complexed with inorganic phosphate
(Pi), which is a strong competitive inhibitor as well as a substrate for the
reverse reaction, has been refined at 2.0 A resolution. Some parts of the
molecular have been retraced, starting from the previous 2.8 A study. The active
site has been modified substantially and is described in this paper. The changes
in the active site region suggest the need to reinterpret earlier spectral data,
and suggestions are made. Also presented are the structures of the
Cd-substituted enzyme complexed with inorganic phosphate at 2.5 A resolution,
and the phosphate-free native enzyme at 2.8 A resolution. At pH 7.5, where the
X-ray data were collected, the Cd-substituted enzyme is predominantly the
covalent phosphoenzyme (E-P) while the native Zn/Mg enzyme exists in
predominantly noncovalent (E.P) form. Implication of these results for the
catalytic mechanism of the enzyme is discussed. APs from other sources are
believed to function in a similar manner.
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